The overall goal of this proposal is to investigate mitochondrial DNA (mtDNA) mutations in prostate cancer and determine the mechanisms by which such mutations protect cells from apoptosis, enhance tumor growth and metastasis and mediate the epithelial response to the bone stromal environment. Our preliminary data indicate that mtDNA mutations are common in clinical prostate cancer specimens and that the introduction of mtDNA mutations in cultured prostate cancer cells decreases apoptosis while increasing tumor formation. With mtDNA mutation there is an approximately 10-fold increase in cellular reactive oxygen production, which may be a mitogenic signal resulting from mtDNA mutation. Furthermore, bone stromal cells are a rich source of ROS under physiologic conditions, a condition that is conducive to 3-dimensional prostate cancer epithelial cell growth, and ROS have been specifically implicated in metastasis. Therefore, because of the clinical observation of mtDNA mutations in prostate cancer and the laboratory association of mtDNA mutations with decreased apoptosis and enhanced tumor growth, we will test the hypothesis that mtDNA mutations enhance prostate cancer epithelial cell growth, tumorigenesis, and metastasis by conferring a survival advantage to the cancer cells, an effect that is enhanced by interactions with bone stromal cells. The testing of this hypothesis involves the following approaches: 1) Introduction of precise mtDNA mutations from clinical prostate cancer into laboratory cell lines by cybrid transfer combined with analysis of growth characteristics, metastatic potential and the interaction with bone stromal cells. 2) We will explore whether reactive oxygen species (ROS) may serve as paracrine mediators of the communication between bone stromal and prostate cancer epithelial cells, using cell culture and chimeric tumors. 3) Evaluation of the mechanisms of altered apoptosis observed in tumors with mtDNA mutations, including changes in the expression of the Bcl-family of proteins.